Background Robot-assisted hand function therapy is pivotal in the rehabilitation of patients with stroke; however, its therapeutic mechanism remains elusive. Currently, research examining the impact of robot-assisted hand function therapy on brain function in patients with stroke is scarce, and there is a lack of studies investigating the correlation between muscle activity and alterations in brain function. Objective This study aimed to investigate the correlation between forearm muscle movement and brain functional activation by employing the synchronized use of functional near-infrared spectroscopy and surface electromyography methods. Moreover, it sought to compare neural activity patterns during different rehabilitation tasks and refine the mechanism of robot-assisted hand function therapy for post-stroke hand function impairments. Methods Stroke patients with hand dysfunction underwent three sessions of robot-assisted hand function therapy within 2 weeks to 3 months of onset. The fNIRS-sEMG synchronous technique was used to observe brain function and forearm muscle activation. Ten participants were randomly assigned to receive mirror, resistance, or passive rehabilitation training. During the intervention, cortical and muscle activation information was obtained using fNIRS and electromyographic signals. The primary outcomes included changes in oxyhemoglobin concentration and root mean square of surface electromyography. Results Compared to the resting state, the Oxy-Hb concentration in the brain regions involved in three rehabilitation tasks with robot-assisted hand function therapy significantly increased (p< 0.05). Mirror therapy significantly enhanced the prefrontal cortex and the superior frontal cortex activation levels. In contrast, resistance therapy significantly promoted the activation of the supplementary motor area and the premotor cortex. Passive rehabilitation tasks showed some activation in the target brain area premotor cortex region. Robot-assisted hand function therapy has shown that forearm muscle movement is closely related to oxygenated hemoglobin concentration activity in specific brain regions during different rehabilitation tasks. Conclusion The simultaneous sEMG-fNIRS study found a significant correlation between muscle movement and brain activity after stroke, which provides an important basis for understanding the treatment mechanism of hand function impairment.